Abstract

The structural characterization of subtilisin mesoscale clusters, which were previously shown to induce supramolecular order in biocatalytic self-assembly of Fmoc-dipeptides, was carried out by synchrotron small-angle X-ray, dynamic, and static light scattering measurements. Subtilisin molecules self-assemble to form supramolecular structures in phosphate buffer solutions. Structural arrangement of subtilisin clusters at 55degC was found to vary systematically with increasing enzyme concentration. Static light scattering measurements showed the cluster structure to be consistent with a fractal-like arrangement, with fractal dimension varying from 1.8 to 2.6 with increasing concentration for low to moderate enzyme concentrations. This was followed by a structural transition around the enzyme concentration of 0.5 mg/mL to more compact structures with significantly slower relaxation dynamics, as evidenced by dynamic light scattering measurements. These concentration-dependent supramolecular enzyme clusters provide tunable templates for biocatalytic self-assembly.